Major histocompatibility complex (MHC) class I-restricted αβ T cells express the CD8αβ heterodimer, which acts as a MHC class I-specific co-receptor. Rats are so far the only species with frequent expression of the CD8αβ by MHC-unrestricted γδ T cells. This study compares CD8αβ expression by splenic rat αβ and γδ T cells and reveals a lineage-specific difference in the control of CD8β expression. After activation in vitro, many γδ T cells, but not αβ T cells, persistently down-modulate the expression of CD8β, but not CD8α, at the RNA level. Down-regulation occurred after stimulation with T-cell receptor (TCR)-specific monoclonal antibody (mAb) and interleukin-2 (IL-2) or CD28-mediated costimulation, and after activation with phorbol 12-myristate 13-acetate (PMA) and ionomycin. Functional differences between modulating and non-modulating cells were not found with respect to interferon-γ (IFN-γ) production and cytolytic activity. The modulation could be indicative for a fundamental difference between αβ and γδ T cells and also limits the use of CD8β as a stable marker of γδ T-cell subsets. Possibly, CD8β modulation provides a mechanism to escape over-stimulation by (auto-)antigens by increasing the threshold of TCR-mediated activation in γδ T cells.